Using cloned cDNA probes, we have isolated the genomic sequences for Alpha and Beta actin, vimentin, a major intermediate filament protein, and GAPDH (glyceraldehyde phosphate dehydrogenase). Preliminary characterization of the actin gene family suggests there are two different Alpha genes and one Beta gene. DNA sequence studies around the 5' proximal intron in the 2 Alpha gene isolates demonstrate complete coding sequence homology yet no homology within the intron. In vivo transcription of the vimentin gene, a single copy gene, yields two distinct mRNA transcripts by Northern analysis. Sequence studies of various vimentin cDNA clones demonstrate the 3' noncoding region of the gene contains 4 polyadenylation sites clustered in pairs approximately 300 base pairs apart. The in vivo mRNA transcripts differ by approximately 300 bp. The cDNA clone containing the poly A sites nearest the coding region hybridizes to both in vivo mRNA transcripts; the cDNA clone containing the more distal polyA sites hybridize only to the larger mRNA transcript. This suggests either polyadenylation cluster is used in vivo. The function of this heterogeneity is unclear at present. The GAP cDNA clone from chicken was used to isolate the homologous genes in yeast. Two of the three different yeast GAP genes were characterized with regard to transcription initiation and termination sites.